Elevated atmospheric CO2 modifies responses to water-stress and flowering of Mediterranean desert truffle mycorrhizal shrubs

Predicted increases in atmospheric concentration of carbon dioxide (CO₂) coupled with increased temperatures and drought are expected to strongly influence the development of most of the plant species in the world, especially in areas with high risk of desertification like the Mediterranean basin. Helianthemum almeriense is an ecologically important Mediterranean shrub with an added interest because it serves as the host for the Terfezia claveryi mycorrhizal fungus, which is a desert truffle with increasingly commercial interest. Although both plant and fungi are known to be well adapted to dry conditions, it is still uncertain how the increase in atmospheric CO₂ will influence them. In this article we have addressed the physiological responses of H. almeriense × T. claveryi mycorrhizal plants to increases in atmospheric CO₂ coupled with drought and high vapor pressure deficit. This work reports one of the few estimations of mesophyll conductance in a drought deciduous Mediterranean shrub and evaluates its role in photosynthesis limitation. High atmospheric CO₂ concentrations help desert truffle mycorrhizal plants to cope with the adverse effects of progressive drought during Mediterranean springs by improving carbon net assimilation, intrinsic water use efficiency and dispersal of the species through increased flowering events.     

Effects of iron chlorosis and iron resupply on leaf xylem architecture, water relations, gas exchange and stomatal performance of field-grown peach (Prunus persica)

There is increasing evidence suggesting that iron (Fe) deficiency induces not only leaf chlorosis and a decline of photosynthesis, but also structural changes in leaf morphology, which might affect the functionality of leaves. In this study, we investigated the effects of Fe deficiency on the water relations of peach ( Prunus persica (L.) Batsch.) leaves and the responses of previously chlorotic leaves to Fe resupply via the root or the leaf. Iron deficiency induced a decline of maximum potential photosystem II (PSII) efficiency (F _V/F _M), of rates of net photosynthesis and transpiration and of water use efficiency. Iron chlorosis was associated with a reduction of leaf xylem vessel size and of leaf hydraulic conductance. In the course of the day, water potentials in chlorotic leaves remained higher (less negative) than in green leaves. In chlorotic leaves, normal stomatal functioning was disturbed, as evidenced by the lack of opening upon withdrawal of external CO₂ and stomatal closure after sudden illumination of previously darkened leaves. We conclude that the Fe deficiency induced limitations of xylem conductivity elicited a water saving strategy, which poses an additional challenge to plant growth on high pH, calcareous soils. Fertilisation with Fe improved photosynthetic performance but the proper xylem structure and water relations of leaves were not fully restored, indicating that Fe must be available at the first stages of leaf growth and development.     

Intraspecific variation in Pinus pinaster PSII photochemical efficiency in response to winter stress and freezing temperatures

As part of a program to select maritime pine (Pinus pinaster Ait.) genotypes for resistance to low winter temperatures, we examined variation in photosystem II activity by chlorophyll fluorescence. Populations and families within populations from contrasting climates were tested during two consecutive winters through two progeny trials, one located at a continental and xeric site and one at a mesic site with Atlantic influence. We also obtained the LT₅₀, or the temperature that causes 50% damage, by controlled freezing and the subsequent analysis of chlorophyll fluorescence in needles and stems that were collected from populations at the continental trial site. P. pinaster showed sensitivity to winter stress at the continental site, during the colder winter. The combination of low temperatures, high solar irradiation and low precipitation caused sustained decreases in maximal photochemical efficiency (F_v/F_m), quantum yield of non-cyclic electron transport (Φ_PSII) and photochemical quenching (qP). The variation in photochemical parameters was larger among families than among populations, and population differences appeared only under the harshest conditions at the continental site. As expected, the environmental effects (winter and site) on the photochemical parameters were much larger than the genotypic effects (population or family). LT₅₀ was closely related to the minimum winter temperatures of the population''s range. The dark-adapted F_v/F_m ratio discriminated clearly between interior and coastal populations.In conclusion, variations in F_v/F_m, Φ_PSII, qP and non-photochemical quenching (NPQ) in response to winter stress were primarily due to the differences between the winter conditions and the sites and secondarily due to the differences among families and their interactions with the environment. Populations from continental climates showed higher frost tolerance (LT₅₀) than coastal populations that typically experience mild winters. Therefore, LT₅₀, as estimated by F_v/F_m, is a reliable indicator of frost tolerance among P. pinaster populations.     

Spatial variation in the fatty acid composition of elaiosomes in an ant-dispersed plant: Differences within and between individuals and populations

Euphorbia characias is a common myrmecochorous plant of the western Mediterranean Basin whose seeds are dispersed by ants following fruit explosion. The variation in elaiosomes’ fatty acid composition of this species was studied at three hierarchical levels (sub-individual, individual and population) in four populations from the Iberian Peninsula. We found that differences in fruit location on the inflorescence do not seem to influence the fatty acid composition of elaiosomes, providing to each propagule an equal chance of being dispersed. However, significant differences in elaiosome fatty acid composition between individuals and populations were found for most of the compounds identified. The content of oleic acid, a key mediator in the ant–seed interaction, differed widely between populations, probably reflecting geographic variations in co-adaptation between plants and their dispersers. The finding that the fatty acid composition of E. characias elaiosomes is distinct from that of the seed itself, but very similar to that of elaiosomes from unrelated species, reinforces the idea of convergent evolution in the chemical composition of these structures.     

Photochemistry, remotely sensed physiological reflectance index and de-epoxidation state of the xanthophyll cycle in Quercus coccifera under intense drought

Quercus coccifera L. is a Mediterranean sclerophyllous shrub with a high capacity to resist intense drought stress. Therefore, it could be used in the study of physiological changes suffered by plants at very low water potentials. A remote sensing sensor was used to measure continuously the physiological reflectance index (PRI; defined as the changes in reflectance at 531 nm with respect to those at 570 nm; PRI = [(R531 − R570)/(R531 + R570)] at canopy level and under field conditions in an artificial carpet of seedlings of Q. coccifera during a drought cycle. Correlations between leaf level-measured chlorophyll fluorescence parameters as well as the de-epoxidation state of the xanthophyll cycle [(A + Z)/(V + A + Z)] and canopy level-measured PRI were reasonably good (R ² = 0.57–0.63, P < 0.01), and quite interesting for water stress remote sensing purposes. The instrument’s temporal resolution allowed us to follow the rapid response of PRI to changing photosynthetic active radiation, and to resolve, in response to cloud-induced changes in light intensity, a fast and a slow PRI component. We report the disappearance of the rapid one under conditions of intense drought in response to a sudden increase in light intensity. The underlying photoprotection mechanisms that Q. coccifera shows in response to intense drought stress periods seem to be related to the existence of a low intrathylakoid lumenal pH at the end of the drought cycle. Under intense drought, these mechanisms allow this species to avoid oxidative damage, which was evidenced by the maintenance of an unaltered photosynthetic pigment composition and constant photosystem II efficiency in the mornings. It is concluded that, contrary to early reports, PRI is a sensible, indirect, non-destructive water stress indicator, even in plants experiencing intense drought. Preliminary results of this work were presented at the 3rd International Workshop on Remote Sensing of Vegetation Fluorescence (February 2007, Florence, Italy).     

Relationship between hydraulic resistance and leaf morphology in broadleaf Quercus species: a new interpretation of leaf lobation

We investigated the relationship between leaf shape and leaf hydraulic resistance in a set of broadleaf Quercus tree species (Q. cerris, Q. frainetto, Q. petraea, Q. pyrenaica, Q. robur, Q. rubra, Q. velutina). Seedlings of all the studied species were grown under uniform environmental conditions. A new high-pressure flowmeter was designed to measure leaf-blade hydraulic resistance. Leaf shape was characterised by the complexity of leaf outline which was regarded as an estimate of leaf lobation. This was done using the box-counting fractal dimension of the leaf silhouette. Leaf hydraulic resistance was negatively related to leaf lobation. It is suggested that the lower hydraulic resistance in deeply lobed leaves may constitute a mechanism for improving water balance under dry atmospheric conditions.     

Kinase-mediated trapping of bi-functional conjugates of paclitaxel or vinblastine with thymidine in cancer cells

In the present work, we explore the possibility of introducing selectivity to existing chemotherapeutics via the design of non-pro-drug, bi-functional molecules comprising a microtubule-binding agent and a substrate for a disease-associated kinase. The design, synthesis, and in vitro biological evaluation of paclitaxel-thymidine and vinblastine-thymidine bi-functional conjugates are reported here. This work provides the first account of 'kinase-mediated trapping' of cancer therapeutics.     

Wettability,Polarity, and Water Absorption of Holm Oak Leaves: Effect of Leaf Side and Age

Plant trichomes play important protective functions and may have a major influence on leaf surface wettability. With the aim of gaining insight into trichome structure, composition, and function in relation to water-plant surface interactions, we analyzed the adaxial and abaxial leaf surface of holm oak (Quercus ilex) as a model. By measuring the leaf water potential 24 h after the deposition of water drops onto abaxial and adaxial surfaces, evidence for water penetration through the upper leaf side was gained in young and mature leaves. The structure and chemical composition of the abaxial (always present) and adaxial (occurring only in young leaves) trichomes were analyzed by various microscopic and analytical procedures. The adaxial surfaces were wettable and had a high degree of water drop adhesion in contrast to the highly unwettable and water-repellent abaxial holm oak leaf sides. The surface free energy and solubility parameter decreased with leaf age, with higher values determined for the adaxial sides. All holm oak leaf trichomes were covered with a cuticle. The abaxial trichomes were composed of 8% soluble waxes, 49% cutin, and 43% polysaccharides. For the adaxial side, it is concluded that trichomes and the scars after trichome shedding contribute to water uptake, while the abaxial leaf side is highly hydrophobic due to its high degree of pubescence and different trichome structure, composition, and density. Results are interpreted in terms of water-plant surface interactions, plant surface physical chemistry, and plant ecophysiology.Plant surfaces have an important protecting function against multiple biotic and abiotic stress factors (Riederer, 2006). They may, for example, limit the attack of insects (Eigenbrode and Jetter, 2002) or pathogenic fungi (Gniwotta et al., 2005; Łaźniewska et al., 2012), avoid damage caused by high intensities of UV and visible radiation (Reicosky and Hanover, 1978; Karabourniotis and Bormann, 1999), help to regulate leaf temperature (Ehleringer and Björkman, 1978; Ripley et al., 1999), and chiefly prevent plant organs from dehydration (Riederer and Schreiber, 2001).The epidermis of plants has been found to have a major degree of physical and chemical variability and may often contain specialized cells such as trichomes or stomata (Roth-Nebelsick et al., 2009; Javelle et al., 2011). Most aerial organs are covered with an extracellular and generally lipid-rich layer named the cuticle, which is typically composed of waxes embedded in (intracuticular waxes) or deposited on (epicuticular waxes) a biopolymer matrix of cutin, forming a network of cross-esterified hydroxy C₁₆ and/or C₁₈ fatty acids, and/or cutan, with variable amounts of polysaccharides and phenolics (Domínguez et al., 2011; Yeats and Rose, 2013). Different nano- and/or microscale levels of plant surface sculpturing have been observed by scanning electron microscopy (SEM), generally in relation to the topography of epicuticular waxes, cuticular folds, and epidermal cells (Koch and Barthlott, 2009). Such surface features together with their chemical composition (Khayet and Fernández, 2012) may lead to a high degree of roughness and hydrophobicity (Koch and Barthlott, 2009; Konrad et al., 2012). The interactions of plant surfaces with water have been addressed in some investigations (Brewer et al., 1991; Brewer and Smith, 1997; Pandey and Nagar, 2003; Hanba et al., 2004; Dietz et al., 2007; Holder, 2007a, 2007b; Fernández et al., 2011, 2014; Roth-Nebelsick et al., 2012; Wen et al., 2012; Urrego-Pereira et al., 2013) and are a topic of growing interest for plant ecophysiology (Helliker and Griffiths, 2007; Aryal and Neuner, 2010; Limm and Dawson, 2010; Kim and Lee, 2011; Berry and Smith, 2012; Berry et al., 2013; Rosado and Holder, 2013; Helliker, 2014). On the other hand, the mechanisms of foliar uptake of water and solutes by plant surfaces are still not fully understood (Fernández and Eichert, 2009; Burkhardt and Hunsche, 2013), but they may play an important ecophysiological role (Limm et al., 2009; Johnstone and Dawson, 2010; Adamec, 2013; Berry et al., 2014).The importance of trichomes and pubescent layers on water drop-plant surface interactions and on the subsequent potential water uptake into the organs has been analyzed in some investigations (Fahn, 1986; Brewer et al., 1991; Grammatikopoulos and Manetas, 1994; Brewer and Smith, 1997; Pierce et al., 2001; Kenzo et al., 2008; Fernández et al., 2011, 2014; Burrows et al., 2013). Trichomes are unicellular or multicellular and glandular or nonglandular appendages, which originate from epidermal cells only and develop outwards on the surface of plant organs (Werker, 2000). Nonglandular trichomes are categorized according to their morphology and exhibit a major variability in size, morphology, and function. On the other hand, glandular trichomes are classified by the secretory materials they excrete, accumulate, or absorb (Johnson, 1975; Werker, 2000; Wagner et al., 2004). Trichomes can be often found in xeromorphic leaves and in young organs (Fahn, 1986; Karabourniotis et al., 1995). The occurrence of protecting leaf trichomes has been also reported for Mediterranean species such as holm oak (Quercus ilex; Karabourniotis et al., 1995, 1998; Morales et al., 2002; Karioti et al., 2011; Camarero et al., 2012). There is limited information about the nature of the surface of trichomes, but they are also covered with a cuticle similarly to other epidermal cell types (Fernández et al., 2011, 2014).In this study and using holm oak as a model, we assessed, for the first time, the leaf surface-water relations of the abaxial (always pubescent) versus the adaxial (only pubescent in developing leaves and for a few months) surface, including their capacity to absorb surface-deposited water drops. Based on membrane science methodologies (Fernández et al., 2011; Khayet and Fernández, 2012) and following a new integrative approach, the chemical, physical, and anatomical properties of holm oak leaf surfaces and trichomes were analyzed, with the aim of addressing the following questions. Are young and mature adaxial and abaxial leaf surfaces capable of absorbing water deposited as drops on to the surfaces? Are young and mature abaxial and adaxial leaf surfaces similar in relation to their wettability, hydrophobicity, polarity, work of adhesion (W_a) for water, solubility parameter (δ), and surface free energy (γ)? What is the physical and chemical nature of the adaxial versus the abaxial trichomes, chiefly in relation to young leaves?     

Relationship between ultrasonic properties and structural changes in the mesophyll during leaf dehydration

The broad-band ultrasonic spectroscopy technique allows the determination of changes in the relative water content (RWC) of leaves with contrasting structural features. Specifically, the standardized frequency associated with the maximum transmittance (f/f(o)) is strongly related to the RWC. This relationship is characterized by the existence of two phases separated by an inflexion point (associated with the turgor loss point). To obtain a better understanding of the strong relationship found between RWC and f/f(o), this work has studied the structural changes experienced by Quercus muehlenbergii leaves during dehydration in terms of ultrasounds measurements, cell wall elasticity, leaf thickness, leaf density, and leaf structure. The results suggest that the decrease found in f/f(o) before the turgor loss point can be attributed to the occurrence of changes in the estimation of the macroscopic effective elastic constant of the leaf (c(33)), mainly associated with changes in the bulk modulus of elasticity of the cell wall (ε). These changes are overriding or compensating for the thickness decreases recorded during this phase. On the other hand, the high degree of cell shrinkage and stretching found in the mesophyll cells during the second phase seem to explain the changes in the acoustic properties of the leaf beyond the turgor loss point. The formation of large intercellular spaces, which increased the irregularity in the acoustic pathway, may explain the increase of the attenuation coefficient of ultrasounds once the turgor loss point threshold is exceeded. The direct measurement of c(33) from ultrasonic measurements would allow a better knowledge of the overall biomechanical properties of the leaf further than those derived from the P-V analysis.     

Radial-growth and wood-anatomical changes in overaged Quercus pyrenaica coppice stands: functional responses in a new Mediterranean landscape

Recent land-use changes in intensively managed forests such as Mediterranean coppice stands might profoundly alter their structure and function. We assessed how the abandonment of traditional management practices in coppice stands, which consisted of short cutting-cycles (10–15 years), has caused overaging (stems are usually much older than when they were coppiced) and altered their wood anatomy and hydraulic architecture. We studied the recent changes of wood anatomy, radial growth, and hydraulic architecture in two stands of Quercus pyrenaica, a transitional Mediterranean oak with ring-porous wood forming coppice stands in W–NW Spain. We selected a xeric and a mesic site because of their contrasting climates and disturbance histories. The xeric site experienced an intense defoliation after the severe 1993–1994 summer drought. The mesic site was thinned in late 1994. We studied the temporal variability in width, vessel number and diameter, and predicted the hydraulic conductivities (K _h) of earlywood and latewood. In the mesic site, we estimated the vulnerability to xylem cavitation of earlywood vessels. Overaging caused a steep decline in latewood production at a cambial age of 14 years., which was close to the customary cutting cycle of Q. pyrenaica. The diameter distribution of vessels was bimodal, and latewood vessels only accounted for 4% of the K _h. Overaging, acting as a predisposing factor in the decline episode, was observed at the xeric site, where most trees did not produce latewood in 1993–1995. At the mesic site, thinned trees formed wider tree-rings, more latewood and multiseriate tree-rings than overaged trees. The growth enhancement remained 8 years after thinning. Most of the hydraulic conductivity in earlywood was lost in a narrow range of potentials, between −2.5 and −3.5 MPa. We have shown how hydraulic conductivity and radial growth are closely related in Q. pyrenaica and how aging modulates this relationship.